This protocol shows that it is possible to cause cartilage damage in animals, larger animals without any surgical intervention or structured destruction of the animal knee. The biggest advantage is that the proves to be repetitive in operators even with no experience, which is difficult to achieve with any surgical method. Begin by mounting the anesthetized rat on the fixation device.
Place the anesthetized rat lying on its belly on the base plate with its right knee attached to a small piece of resin with a concave groove. Place the right hind limb in the hip extension, knee flexion, and ankle extension positions with the knee flexed at approximately 140 degrees. Accommodate the heel of the rat on the wedge-shaped groove on the movable fixture.
Move the fixation device to the stress and tensile testing instrument and ensure that there is no contact with the load cell. Open the stress and tensile testing instrument control software and click on the calibration button. After calibration, carefully attach the top of the frame to the load cell.
Turn on the rotary knob on the movable main operational panel slowly until the preload reaches five Newtons to keep the knee joint closely attached to the frame. On the main menu, click on create a new method and system label. Set the test mode to cycle and the test type to compression.
Click on the sensor label and select the test tab to check that the limit is within 60 Newtons. Then select the stroke tab and check that the limit is within 500 millimeters. Under the testing control label, select origin of growth to start the main program with 0.3%or full scale.
Set the stroke speed in control in the first and third sections to one millimeter per second, then set the maximum testing force in the second section to 20 Newtons and the minimum testing force in the fourth section to five Newtons. Set the duration of whole to 0.5 seconds for the peak load and 10 seconds for the minimum load. In the preload tab at the bottom of the page, ensure that on is checked, the speed of deflection removal is set to 100 millimeters per minute, and the maximum force is five Newtons.
Set the material as metal in the specimen label. In the main menu under the select method and test section, select the method that was just built and click on start to begin the test. Set the number of cycles to 60.
Return the rat to its cage after loading and maintain a 12-12 hour light dark schedule in a cage with sufficient space and food ad libitum until it is time for analysis. The short term changes in chondrocyte viability in samples subjected to 20 Newtons of cyclic loading was studied. The number of dead chondrocytes increased at 12 hours post-trauma.
Conversely, the number of living chondrocytes continued to decrease. The articular cartilage of the rat knees that underwent 20 Newtons of dynamic loading was damaged and one focal lesion zone was confirmed in the lateral femoral condyle in all the samples. The lesion size did not progressively increase during the eight week observational period.
The border corresponding to the interface of the lesion and the unaffected cartilage could be observed in the affected area. The most important thing is the proper settings of the testing force and the number of cycles. With this technique, one can control mechanical loading to the articular coverage of blood, paving the way to explore how mechanical loading affect the development or prevention of osteoarthritis.
